Abstract

Farnesyl diphosphate synthase (FPPS) is a crucial protein in terpenoid production. However, its industrial application is limited owing to its low solubility in Escherichia coli. In this study, we focused on ispA encoding FPPS and designed a fusion expression system to reduce inclusion body (IB) formation. Among the chosen fusion tags, the GB1-domain (GB1) exhibited the highest ability to solubilize the recombinant protein. Increased rare tRNA abundance not only improved the GB1-FPPS yield but also increased its soluble level. A “one-step” method for the acquisition of soluble FPPS was also considered. By combining GB1-FPPS expression and Tobacco Etch Virus protease (TEVp) cleavage in vivo, a controllable GB1-FPPS “self-cleavage” system was constructed. Overall, this study provides an efficient approach for obtaining soluble forms of FPPS, which show great potential for use in the soluble expression of other homologous diphosphate synthase.

摘要

法尼基二磷酸合酶 (FPPS) 是萜类化合物生产中的关键蛋白质。然而，由于其在大肠杆菌中的溶解度较低，其工业应用受到限制。在本研究中，我们重点关注编码 FPPS 的ispA，并设计了融合表达系统来减少包涵体 (IB) 的形成。在所选的融合标签中，GB1 结构域 (GB1) 表现出最高的溶解重组蛋白的能力。稀有 tRNA 丰度的增加不仅提高了 GB1-FPPS 的产量，还提高了其可溶水平。还考虑了获取可溶性 FPPS 的“一步”方法。通过将GB1-FPPS表达与烟草蚀纹病毒蛋白酶（TEVp）体内裂解相结合，构建了可控的GB1-FPPS“自裂解”系统。总体而言，这项研究提供了一种获得可溶性形式 FPPS 的有效方法，该方法在其他同源二磷酸合酶的可溶性表达中显示出巨大的潜力。